Contractile Responses in Arteries Subjected to Hypertensive Pressure in Seven-Day Organ Culture

Early stage changes in hypertensive arteries have a significant effect on the long-term adaptation of the arteries. Compared to the long-term adaptation, little is known about the early dimensional and functional changes in hypertensive arteries in the first few days of hypertension. To study the early stage changes in hypertensive arteries, porcine common carotid arteries were cultured for seven days in a simplified ex vivo artery organ culture system with pulsatile flow under hypertensive (200±30 mm Hg) or normotensive (100±20 mm Hg) pressure conditions while maintaining a physiological mean wall shear stress of 15 dyn/cm².Vessel viability was demonstrated by contractile diameter responses to norepinephrine (NE), carbachol (CCh), and sodium nitroprusside (SNP) as well as staining for mitochondrial activity and cell apoptosis/necrosis. The results show that arteries demonstrated strong contractile responses to NE, CCh, and SNP, basal tone, and viable mitochondria in the organ culture system for seven days. Hypertensive arteries demonstrated a stronger contractile response than normotensive arteries (p < 0.05). Diameter enlargement was observed in hypertensive arteries as compared to arteries cultured under normotensive conditions. In conclusion, the pulsatile culture system can maintain arteries viable with active vasomotion tone for up to seven days. Hypertensive pressure causes arterial adaptation by significantly increasing arterial diameter and contractile response within the first seven days. © 2001 Biomedical Engineering Society. PAC01: 8719Uv, 8719Rr, 8780Rb     

Sustained Axial Loading Lengthens Arteries in Organ Culture

Although it has been recognized for many years that arteries in vivo exist under significant axial strain, studies of the adaptation of arteries to elevated axial strain have only recently been conducted. To determine the effects of sustained elevation of axial loading on arterial structure and function, axial stresses of 250 kPa or greater were applied to porcine common carotid arteries maintained in a perfusion organ culture system for 7 days at physiologic pressure and flow conditions. Our results demonstrated that axial stretch could lead to an increase in unloaded length that was proportional to the axial stretch ratio (stretched length divided by unloaded length) when the axial stretch ratio was above a threshold value of 2.14. Below this threshold, no significant length change occurred. Above this threshold, a significant increase in unloaded length (13 ± %,) and the number of smooth muscle cell nuclei (20 ± 7%) was observed. Permanent length change was associated with a significant decrease in axial stiffness, and the maximum elongation achieved was limited by rupture of the arterial wall. All tested arteries demonstrated good viability and strong vasomotor responses. These results show that arteries in organ culture can elongate under sustained axial loading.     

Pronephros and spleen cells from rainbow trout (Oncorhynchus mykiss) in vitro: Growth factors produced under the influence of mitogens

Supernatants or conditioned media (CM) were produced by rainbow trout pronephros cells (PNC) and spleen cell cultures (1×10⁶/ml), by addition of 20 g/ml phythaemagglutinin (PHA), 5 ng/ml 12-O-tetradecanoyl-phorbol-13-acetate (PHA) (for 2h), PHA together with PMA, 10% horse serum (HS), or 100 g/ml concanavalin A (ConA) after a culture of 7 days. Only PNC were effective growth factor producers. The effect of different concentrations of CM (5%–28%) on cell number was tested after a cultivation period of 14 days. PNC at a concentration of 1×10⁶/ml were cultured with various concentrations of CM and the proliferation was tested by the XTT-test (testing the dehydrogenase activity of the cells by formation of a formazan) after 10 days. CM produced by cells with PHA, PHA and PMA and HS increased the proliferation in a concentration-dependent manner. CM produced with PMA alone was effective in the XTT-test. There was no synergistic or additive effect of PMA with PHA. CM produced with ConA had no effect, although in the XTT-test a strong proliferation of PNC in presence of 100 g/ml ConA was observed. In a semisolid culture with collagen, CM treatment resulted in prevention of cell death and an increase in cell size but did not induce proliferation. All CM obtained from spleen cells had no effect. Stimulation of spleen cells by CM could be seen only in the XTT-test. PHA and HS trigger the PNC to release growth factors in vitro which stimulate cell growth and/or prevent cell death.     

Role of Rac and Rho-GDI Alpha in the Frequency-dependent Expression of h1-calponin in Vascular Smooth Muscle Cells under Cyclic Mechanical Strain

Phenotype transformation of vascular smooth muscle cells (VSMCs) has been reported to be directly influenced by the frequency of mechanical strain. This study explored the effects of different frequencies of mechanical strain on expression of phenotype marker h1-calponin and the possible mechanism. VSMCs were subjected to cyclic strains of 10% elongation at 1 and 2 Hz for 24 h by using a Flexercell strain unit. The protein expression of h1-calponin was assessed by Western blotting and the possible protein kinases involved were evaluated by their specific inhibitor or targeted siRNA ‘knock-down.’ The results showed that cyclic strains modulated the expressions of h1-calponin, phospho-p38, Rac and Rho-guanine nucleotide dissociation inhibitor alpha (Rho-GDIα) in nonlinear frequency-dependent manners. This nonlinear frequency-dependent change of h1-calponin expression could be blocked by a specific p38 inhibitor, SB202190. The changed expression of phospho-p38 induced by the frequencies of cyclic strain was reversed by targeted siRNA ‘knock-down’ of Rac, while enhanced by targeted siRNA ‘knock-down’ of Rho-GDIα. These results suggest that the frequency-dependent expression of h1-calponin under cyclic strain is mediated at least partly by the regulation of Rac and Rho-GDIα expression on the activation of p38 pathway. Ming-Juan Qu and Bo Liu contributed equally to this work.